Nerve Growth Factor (NGF) has been shown to have two sites of action in the stimulation of neurite outgrowth from chick embryo dorsal root ganglia in vitro. In conjunction with an unknown factor found in rooster plasma but not rooster serum, NGF will stimulate neurite outgrowth in a manner not dependent on de novo RNA synthesis but which requires protein synthesis. Ganglia cultured in media devoid of the plasma, however, require both RNA and protein synthesis to produce neurite outgrowth in response to NGF. This process of neurite outgrowth can be temporarily inhibited by Ca ions applied externally in the medium but the ganglia eventually overcome this inhibition and neurites grow out. Attempts will be made to isolate and characterize the rooster plasma factor which works in conjunction with NGF. Initially, plasma will be fractionated by gel filtration and fractions of different molecular weight material will be assayed for activity by using rooster serum in the ganglial culture with actinomysin D and the fraction to be tested. The mechanism by which Ca ions inhibits neurite outgrowth will also be explored with emphasis being placed on determining how NGF regulates the Ca ions levels. Initial studies will be done to determine the intracellular Ca ions concentration before and after NGF treatment to see if levels are controlled by the plasma membrane Ca ions pump or by a sequestering mechanism in the cell. The rates of synthesis and degradation of both brain tubrilin and actin will be measured during different periods of chick embryo brain development. The regulation of tubrilin levels will be studied by attempting to isolate and characterize the tubrilin mRNA and determine its rate of translation and turnover in brain. Finally, the functional relationships between tubrilin, actin and other proteins of the myofibril identified in brain will be explored by assaying for changes in ATPase activity of brain actomyosin or dynein in the presence of the other proteins.